T lymphocytes undergo two selection processes during development in the thymus: tolerance and positive selection. This grant proposes to study these developmental events by using transgenic mice which express T cell antigen receptors. To a large extent, the transgene encoded receptor replaces endogenously encoded receptors in these animals, allowing one to study development in a large population of T cells with identical specificity. The murine T cell response to pigeon cytochrome c and class II major histocompatibility complex encoded (MHC) molecules will be used as a model system. Experiments will address how the structure of the T cell antigen receptor influences the recognition of MHC molecules during T cell maturation in the thymus. Two separate lines of transgenic animals which express antigen receptors that differ by only a single amino acid will be analyzed. This single amino acid difference results in a change in the pattern of MHC restriction in vitro. Whether this region of the T cell antigen receptor also alters the fine specificity of MHC recognition during positive selection in the thymus will be determined by analyzing the development of T cells in transgenic mice of varied MHC haplotypes. Transgenic mice which express these T cell antigen receptors will also be used to study the mechanisms of tolerance to self-antigens. Two approaches will be used. First, neonatal transgenic mice will be rendered tolerant to pigeon cytochrome c by repeated injection of the antigen. The maturation of T cells in these animals will be analyzed in order to determine the mechanism of tolerance to this soluble antigen. Second, gene constructs encoding a secreted or cytoplasmic form of a fragment of pigeon cytochrome c will be expressed in vitro and in transgenic mice. These experiments will address how the cellular localization of a self-antigen influences its ability to induce tolerance in class II MHC restricted T cells. Double transgenic mice which express a T cell antigen receptor and its specific antigen will also be produced. These mice will allow a careful analysis of the mechanism of tolerance induction to the transgene encoded antigens. The issues concerning T cell development that are addressed by this proposal relate to how a mature T cell repertoire is selected, both by elimination of auto-destructive specificities and by positive selection for self recognition. Certainly these developmental events are central to understanding many facets of autoimmune disease and MHC linked disease susceptibility.